The development and application of biological models for evaluation of direct nose-to-brain drug delivery systems

Mistry, Alpesh (2009) The development and application of biological models for evaluation of direct nose-to-brain drug delivery systems. PhD thesis, University of Nottingham.

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Abstract

The olfactory neuroepithelium is the only part of the central nervous system that is exposed directly to the external environment. Therefore, it is the only non-invasive drug delivery route to the brain. Surface modification of PS nanoparticles with chitosan C-PS), polysorbate 80 (P80-PS) and polysorbate 80+FCS (P80-FCS-PS) changed the toxicity and distribution of these nanoparticles in olfactory mucosae. In addition, a reduction in nanoparticle diameter from 200nm to 20nm increased nanoparticle mucosal penetration and possibly also their cellular toxicity.

In vitro vertical Franz diffusion chamber and in vivo mouse models were adapted to investigate the transport of nanoparticles via the olfactory system. For the in vitro model, preliminary studies found that olfactory epithelium lined the caudal portion of the dorsal nasal turbinate in the porcine nasal cavity. To ensure the scientific validity of the diffusion chamber studies, it was necessary to prove that the experimental procedures themselves (without the addition of nanoparticles) had no effect on the mounted tissue. Therefore, viability and cellular morphology of the dissected olfactory epithelia were assessed prior to application of nanoparticles to tissues. Alamar Blueâ„¢ viability and histological findings showed that the diffusion chamber experiment did not affect the olfactory tissue when compared to samples that were not mounted on the apparatus. Citrate buffer (pH6.0) had significantly reduced the viability (PD, Isc and Alamar Blueâ„¢) of the porcine olfactory epithelium compared to SNS buffer (pH7.4) but it did not kill it. Citrate buffer may have depleted the mucosal pH gradient in the epithelium. Overall both SNS buffered and citrate buffered porcine olfactory epithelia were suitable for nanoparticle transport studies in the vertical Franz diffusion cell.

The in vitro and in vivo biological models showed surface modification had changed the distribution of nanoparticles within the epithelia. There was good agreement between particle losses from donor chamber, fluorescence microscopy images and stereology results that C-PS particles adhered to extracellular mucus to a greater extent compared to PS and P80-FCS-PS. P80-PS nanoparticles were taken into the nasal epithelial cells to a greater extent than C-PS. Nanoparticles were not transported to the receiver chamber in vitro or the olfactory bulbs in vivo. The size of the nanoparticles was also important. Fluorescence microscopy and stereology showed that greater numbers of 100nm PS and 100nm P80-FCS-PS were taken up into mouse olfactory epithelial cells compared to 200nm diameter equivalents. Larger particles may not have penetrated mucus as effectively as smaller ones.

Bright field microscopy images of olfactory epithelia dismounted from the diffusion chamber apparatus after transport study with C-PS nanoparticles showed that these particles caused the greatest amount of cellular damage compared to PS, P80-PS and P80-FCS-PS systems. Greater damage was observed for progressively smaller particles. For example, 20nm C-PS may have accessed subcellular organelles such as mitochondria to cause cell death by oxidative stress. However, similar findings in the mouse model were not observed. It was hypothesised that, unlike the in vitro model, the mouse model may have been able to maintain a pH gradient across the mucous layer by neutralising the acidity from the citrate buffer using blood borne HCO3- ions. This would protect the epithelial cells by causing C-PS to aggregate in the mucus thereby preventing them from accessing the epithelial cells.

Item Type:Thesis (PhD)
Supervisors:Stolnik-Trenkic, S.
Uncontrolled Keywords:Chitosan, Polysorbate 80, Diffusion Chamber, Toxicity, Olfactory
Faculties/Schools:UK Campuses > Faculty of Science > School of Pharmacy
ID Code:654
Deposited By:Alpesh Mistry
Deposited On:23 Jun 2010 15:38
Last Modified:23 Jun 2010 15:38

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